Although a natural infection by Bacillus anthracis is rare, anthrax spores can be effectively distributed as biological weapons in many different means, such as the surface contamination of the U.S. mail occurred in the post September 11 attacks. Thus, effective approaches for anthrax prophylaxis are needed. The current anthrax vaccine in the U.S., the anthrax vaccine adsorbed (AVA), has many drawbacks including the requirement for multiple injections with subsequent annual boosters. New anthrax vaccine developments have been focused on the production of a vaccine based on the protective antigen (PA) component of the anthrax toxin. Topical non-invasive immunization onto the skin with a PA-encoding plasmid DNA vaccine is more convenient, patient friendly, cost-effective, and thus, may be more feasible for the immunization of a large population of people in case of an emergency. However, the immune response from this route of immunization is often rather weak. Our own preliminary studies strongly suggest that a microemulsion-based DNA vaccine carrier developed by us significantly enhanced the immune response from DNA vaccines, including a PA63 gene encoding DNA vaccine, when topically, non-invasively applied onto the skin of mice. Thus, this microemulsion could potentially be used to topically administer an anthrax DNA vaccine in humans. However, there is a critical need to identify the mechanisms of immune induction/enhancement by this DNA vaccine-incorporated microemulsion and to validate the efficacy of the anti-PA immune response induced before considering such a vaccine as a prophylactic possibility for the prevention of an anthrax infection in humans. Our long-term objective is to develop an efficacious, non-invasive, topical anthrax vaccine. The specific aims in this particular application are to identify the mechanisms involved in the immune induction by this DNA vaccine-incorporated microemulsion and to evaluate the activity of the anti-PA immune response induced in a mouse model. The central hypothesis is that the PA-encoding plasmid DNA-incorporated microemulsion, when topically applied onto the skin, will elicit strong anti-PA immune responses by enhancing the uptake and expression of the encoding PA gene in hair follicles in mice. The successful completion of this project is expected to provide a scientifically sound foundation for the use of this microemulsion-based DNA vaccine in human clinical trials. An effective topical anthrax vaccine will contribute to the protection and improvement of public health. Moreover, the elucidation of the mechanism of immune induction by this vaccine and the demonstration of its efficacy will enable the application of this microemulsion in developing vaccines for other human and animal diseases. [unreadable] [unreadable] [unreadable]